Heterozygous germline BLM mutations increase susceptibility to asbestos and mesothelioma

Rare biallelic BLM gene mutations cause Bloom syndrome. Whether BLM heterozygous germline mutations (BLM^+/−) cause human cancer remains unclear. We sequenced the germline DNA of 155 mesothelioma patients (33 familial and 122 sporadic). We found 2 deleterious germline BLM^+/− mutations within 2 of 33 families with multiple cases of mesothelioma, one from Turkey (c.569_570del; p.R191Kfs*4) and one from the United States (c.968A>G; p.K323R). Some of the relatives who inherited these mutations developed mesothelioma, while none with nonmutated BLM were affected. Furthermore, among 122 patients with sporadic mesothelioma treated at the US National Cancer Institute, 5 carried pathogenic germline BLM^+/− mutations. Therefore, 7 of 155 apparently unrelated mesothelioma patients carried BLM^+/− mutations, significantly higher (P = 6.7E-10) than the expected frequency in a general, unrelated population from the gnomAD database, and 2 of 7 carried the same missense pathogenic mutation c.968A>G (P = 0.0017 given a 0.00039 allele frequency). Experiments in primary mesothelial cells from Blm^+/− mice and in primary human mesothelial cells in which we silenced BLM revealed that reduced BLM levels promote genomic instability while protecting from cell death and promoted TNF-α release. Blm^+/− mice injected intraperitoneally with asbestos had higher levels of proinflammatory M1 macrophages and of TNF-α, IL-1β, IL-3, IL-10, and IL-12 in the peritoneal lavage, findings linked to asbestos carcinogenesis. Blm^+/− mice exposed to asbestos had a significantly shorter survival and higher incidence of mesothelioma compared to controls. We propose that germline BLM^+/− mutations increase the susceptibility to asbestos carcinogenesis, enhancing the risk of developing mesothelioma.

In the United States, the incidence rate of mesothelioma varies between fewer than one case per 100,000 persons in states with no asbestos industry to two to three cases per 100,000 persons in states with an asbestos industry (1, 2). Asbestos causes DNA damage and apoptosis (3) and promotes a chronic inflammatory reaction that supports the emergence of malignant cells (4). Fortunately, only a small fraction of exposed individuals develop mesothelioma; for example, 4.6% of deaths in miners who worked in asbestos mines for over 10 y were caused by mesothelioma (1). Therefore, multiple cases of mesothelioma in the same family are rare and suggest genetic predisposition (5). In 2001, we discovered that susceptibility to mesothelioma was transmitted in a Mendelian fashion across multiple generations in some Turkish families exposed to the carcinogenic fiber erionite, pointing to gene × environment interaction (G×E) as the cause (6). In 2011, we discovered that carriers of heterozygous germline BRCA1-associated protein–1 (BAP1) mutations (BAP1^+/−) developed mesothelioma and uveal melanoma (5), findings expanded and confirmed by us and by multiple research teams (reviewed in refs. 1, 7, 8). Moreover, heterozygous germline Bap1 mutations (Bap1^+/−) significantly increased susceptibility to asbestos-induced mesothelioma in mice (9, 10), evidence of G×E. Reduced BAP1 levels impair DNA repair (11) as well as different forms of cell death (3, 12) and induce metabolic alterations (13–15) that together favor cancer development and growth.Recent studies revealed that mesothelioma may also develop among carriers of germline mutations of additional tumor-suppressor genes that cause well-defined cancer syndromes, including MLH1 and MLH3 (Lynch syndrome), TP53 (Li–Fraumeni syndrome), and BRCA1-2 (Breast and Ovarian Cancer syndrome) (16, 17). When all germline mutations are combined, it has been estimated that about 12% of mesotheliomas occur in carriers of heterozygous germline mutations of BAP1, the most frequent mutation among patients with mesothelioma, or of other tumor suppressors. Some of these mutations may sensitize the host to asbestos carcinogenesis, according to a G×E scenario (17). Thus, presently, mesothelioma is considered an ideal model to study G×E in cancer (17). As part of the Healthy Nevada Project (HNP), we are studying G×E in northern Nevada, a region with an unusually high risk of exposure to carcinogenic minerals and arsenic, which may be related to the high cancer rates in this region (18). We are investigating genetic variants that may increase cancer risk upon exposure to carcinogens to implement preventive strategies.Biallelic mutations of the Bloom syndrome gene (BLM) cause Bloom syndrome, an autosomal-recessive tumor predisposition syndrome characterized by pre- and postnatal growth deficiency, photosensitivity, type 2 diabetes, and greatly increased risk of developing various types of cancers. BLM is a RecQ helicase enzyme that modulates DNA replication and repair of DNA damage by homologous recombination (19). In patients affected by Bloom syndrome, the absence of the BLM protein causes chromosomal instability, increased number of sister chromatid exchanges, and increased numbers of micronuclei (20–22). In addition, BLM is required for p53-mediated apoptosis (23), a process critical to eliminate cells that have accumulated DNA damage. Impaired DNA repair together with altered apoptosis resulted in increased cancer incidence (17, 24). Of course, inactivating germline BLM heterozygous (BLM^+/−) mutations are much more common than biallelic BLM (BLM^−/−) mutations, with an estimated frequency in the general population of 1 in 900 based on data from the Exome Aggregation Consortium (25). BLM^+/− mutation carriers do not show an obvious phenotype; however, some studies have suggested that carriers of these mutations may have an increased cancer risk (17, 24). Mice carrying Blm^+/− mutations are prone to develop a higher rate of malignancies in the presence of contributing factors, such as concurrent heterozygous mutations of the adenomatous polyposis coli (Apc) gene, or upon infection with murine leukemia virus (26). However, in studies in which Blm^+/− mice were crossed with tuberous sclerosis 1-deficient (Tsc1^+/−) mice that are predisposed to renal cystadenomas and carcinomas, Wilson et al. found that Tsc1^+/− Blm^+/− mice did not show significantly more renal cell carcinomas compared with Tsc1^+/− Blm^WT mice (27). In humans, a large study involving 1,244 patients with colon cancer and 1,839 controls of Ashkenazi Jewish ancestry, in which BLM^+/− frequency is as high as 1 in 100 individuals (28), suggested that carriers of germline BLM^+/− mutations might have a twofold increase in colorectal cancer (CRC) (29). A smaller study did not confirm these results, but reported a trend of increasing incidence of adenomas—premalignant lesions—among BLM^+/− mutation carriers (30). In addition, BLM^+/− mutations were found overrepresented among early-onset (<45 y old) CRC patients (25). Other studies associated BLM^+/− mutations to an increased risk of breast (31, 32) and prostate cancer (33), but the low power of these studies hampered definite conclusions. In summary, it appears possible that BLM^+/− mutations may increase cancer risk in the presence of contributing factors.     

Organization of the chemokine gene cluster on human chromosome 17q11.2 containing the genes for CC chemokine MPIF-1, HCC-2, HCC-1, LEC, and RANTES.

To understand the organization of the human CC chemokine gene cluster on chromosome 17q11.2, we determined the nucleotide sequence of a region 181 kb long containing five CC chemokine genes, MPIF-1 (SCYA23), HCC-2 (SCYA15), HCC-1 (SCYA14), LEC (SCYA16), and RANTES (SCYA5), by the random shot-gun method. The four CC chemokine genes, MPIF-1, HCC-2, HCC-1, and LEC, are clustered within a region 40 kb long, whereas the RANTES gene is located approximately 10 kb apart from the four chemokine gene minicluster. These chemokine genes are arranged in the same orientation, and their sizes are relatively long, 3.1 (HCC-1)-8.8 kb (RANTES) when compared with other CC chemokine genes, such as MIP-1alpha/LD78alpha (SCYA3) (1.9 kb) and MCP-1 (SCYA2) (1.5 kb). In contrast to most other human CC chemokine genes that consist of three exons, the MPIF-1 and HCC-2 genes, separated by 12 kb, have four exons. When the nucleotide sequences of the MPIF-1 and HCC-2 genes are compared, they are well conserved, including introns and flanking sequences, except for the middle region of the long first intron, indicating that they have been generated recently in evolutionary terms by duplication. In addition to the CC chemokine genes, more than 30 exons are identified in the sequenced region by similarity search against expressed sequence tags (ESTs) and also by the gene prediction program GenScan. This indicates that the chemokine cluster sequenced in this study is a gene-rich region in the human genome.     

Stimulation of nonspecific resistance to infection induced by muramyl dipeptide analogs substituted in the gamma-carboxyl group and evaluation of N alpha-muramyl dipeptide-N epsilon-stearoyllysine.

Stimulation of resistance to infection induced by the analogs of muramyl dipeptide (MDP) having substituted functions in the gamma-carboxyl group of D-isoglutamyl residue was examined in experimental Escherichia coli infections in mice. An MDP analog which is an efficient strengthener of resistance to infection, N alpha-MDP-N epsilon-stearoyllysine [MDP-Lys(L18)], was selected through the comparative assessment of a number of compounds in three categories: (i) gamma-alkylamides, (ii) gamma-esters, and (iii) N alpha-MDP-N epsilon-acyllysine derivatives. Furthermore, the antiinfectious activity of MDP-Lys(L18) was evaluated bacteriologically in comparison with that of MDP. The effect of MDP-Lys(L18) on the susceptibility of mice to infections with various species of microorganisms was studied. Protective activity was greatest against E. coli and staphylococcal infections, considerable against Pseudomonas and Candida infections, and least against Klebsiella infection. The effects of bacterial inoculum size and MDP treatment timing, dose, and route of administration on protective activity were studied. The efficacy of MDP-Lys(L18) in protection tests was demonstrated for all administration routes, even the oral. Its high potency was confirmed by the smaller influence of inoculum size and particularly small value of the minimum dosage required for inducing protective activity. A decrease in bacterial survival was observed in the blood and organs of mice treated with the analog and infected with E. coli. The following two useful effects were obtained: the synergistic effect of glycopeptide and chemotherapeutic agents and the stimulation of resistance to infection in animals immunocompromised by cyclophosphamide treatment.     

Does supplementation of contrast MR imaging with thallium-201 brain SPECT improve differentiation between benign and malignant ring-like contrast-enhanced cerebral lesions?

Objective To determine whether thallium-201 (²⁰¹Tl) brain single photon emission computed tomography (SPECT) could supplement magnetic resonance (MR) imaging diagnostic information by visual comparison of two separate data sets from patients with ring-like contrast-enhanced cerebral lesions. Methods A combination of MR imaging and ²⁰¹Tl brain SPECT sets obtained from 13 patients (10 men, 3 women) ranging in age from 26 years to 86 years (mean 61.0 years) were retrospectively reviewed. A total of 12 patients had a solitary lesion, and the others had multiple lesions. All but two intracranial foci were pathologically confirmed. The final diagnoses were six glioblastomas, two cerebral metastases from lung cancer, and one each of abscess, resolving hematoma, primary central nervous system lymphoma, toxoplasmosis, and radiation necrosis. The two separate image formats (MR images and SPECT) were shown to ten readers with practical experience. All of the MR images for each patient were shown to each reader first. After interpreting them, the readers were shown the SPECT images. Images were scored in terms of how benign or malignant the foci were on a 5-point scale from “definitely benign” to “definitely malignant.” Results The improvement in the performance of all ten readers was from 67.7% to 93.8% in mean accuracy (P = 0.0028) and from 0.730 to 0.971 in mean Az value (P = 0.0069) after they were shown the ²⁰¹Tl brain SPECT images. Conclusions ²⁰¹Tl brain SPECT should substantially increase confidence in the diagnosis of intracranial lesions with ring-like contrast enhancement when MR imaging does not permit differentiation between benign and malignant disease.     

Contrast-enhanced ultrasonography in the diagnosis of solid renal tumors.

OBJECTIVE: The purpose of this study was to evaluate the usefulness of contrast-enhanced ultrasonography (CEUS) in the diagnosis of solid renal tumors. METHODS: Twenty-nine patients with solid tumors detected on gray scale ultrasonography underwent resection for suspected renal malignancy. Findings of arterial phase contrast computed tomography (CT) and CEUS were compared for each diagnosis. RESULTS: Histopathologic examination of resected lesions showed malignancy in 26 patients (clear cell carcinoma, n = 18; papillary renal cell carcinoma, n = 6; collecting duct carcinoma, n = 1; and infiltrative urothelial carcinoma, n = 1) and benign tumors in 3 patients (oncocytoma, n = 2; and angiomyolipoma, n = 1). Contrast CT failed to show tumor blood flow in 5 of 29 patients, whereas CEUS showed this in all patients. Positive predictive values of CEUS and contrast CT in the diagnosis of renal malignancy were 100% and 82.8%, respectively. Among clear cell carcinomas, hypervascularity was observed on contrast CT in 16 of 18 patients and on CEUS in 17 of 18 patients. On the basis of hypervascularity, diagnostic sensitivity values for clear cell carcinoma were 94.4% for CEUS and 88.9% for contrast CT, whereas specificity values were 45.5% for CEUS and 72.7% for contrast CT. Among papillary cell carcinomas, contrast CT showed avascular lesions in 4 of 6 patients. However, CEUS showed blood flow in these lesions, leading to diagnosis of hypovascular renal tumors. CONCLUSIONS: Contrast-enhanced ultrasonography was more sensitive for detecting slight tumor blood flow than contrast CT and was useful in preoperatively diagnosing malignant hypovascular renal tumors but was less so for hypervascular renal tumors.     

Granulocyte adsorptive apheresis for pediatric patients with ulcerative colitis

Granulocytapheresis (GCAP) has produced efficacy in adult patients with ulcerative colitis (UC) by adsorbing activated granulocytes and monocytes/macrophages. We retrospectively investigated efficacy and safety of GCAP in pediatric patients with active UC. Twelve steroid-refractory children (12.2±3.1 years old) were treated with GCAP, one session/week for 5–10 consecutive weeks. In 8 patients, clinical symptoms improved after two GCAP sessions. Normal body temperature, stool frequency, and disappearance of blood in stool were seen after 24.3±11.5 days. The endoscopic grade improved from 2.6±0.3 to 0.4±0.2. One patient who initially responded, developed bloody diarrhea later and 2 cases remained unchanged. The dose of steroid was tapered during GCAP therapy by 50%. No serious adverse effects were noted. Four of 8 cases relapsed 3.5 ± 2.2 months after the last GCAP while on maintenance therapy, the other 4 were in remission up to 22.8±18.1 months. In conclusion, GCAP appears to be effective and well tolerated in children with steroid-refractory UC.     

Peel-off resection of the pituitary gland for functional pituitary adenomas: pathological significance and impact on pituitary function

Purpose

Functional pituitary adenomas (FPAs) lacking a well-defined pseudocapsule can invade the adjacent pituitary gland. In such situations, peel-off resection of the adjacent pituitary gland after selective adenomectomy might lead to complete tumor removal, resulting in optimal endocrinological outcomes. Here, we present the significance of peel-off resection of the pituitary gland in patients with FPA in whom complete extracapsular tumor removal cannot be achieved.

Methods

We performed a retrospective review of 21 patients with FPA who underwent transsphenoidal surgery (TSS). After selective adenomectomy, peel-off resection of the adjacent pituitary gland was performed in 13 patients because complete extracapsular resection could not be achieved, while peel-off resection was not performed in the remaining 8 patients because complete extracapsular resection was accomplished. The clinical outcomes of these groups were compared. The pituitary tissues obtained by peel-off resection were pathologically examined for tumor cells.

Results

Early postoperative biochemical remission was achieved in 20 patients (95.2%). Anterior pituitary functions were not aggravated postoperatively in any patient: however, transient diabetes insipidus (DI) occurred in 2 patients. There were no statistically significant differences in the clinical outcomes of the two groups. A pseudocapsule was pathologically detected in the adjacent anterior pituitary even in patients in whom no pseudocapsule was intraoperatively detected. Tumor cells were pathologically detected in 7 (58.3%) of 12 pituitary tissues examined.

Conclusions

Peel-off resection of the pituitary gland, which can remove a small tumor cell remnant in the adjacent pituitary, might maximize the effectiveness of TSS with minimal impact on postoperative pituitary function.